Reverse electron flow-induced ROS production is attenuated by activation of mitochondrial Ca²⁺-sensitive K⁺ channels

Abstract

Heinen A, Aldakkak M, Stowe DF, Rhodes SS, Riess ML, Varadarajan SG, Camara AK. Reverse electron flow-induced ROS production is attenuated by activation of mitochondrial Ca 2ϩ -sensitive K ϩ channels. Am J Physiol Heart Circ Physiol 293: H1400-H1407, 2007. First published May 18, 2007; doi:10.1152/ajpheart.00198.2007.-Mitochondria generate reactive oxygen species (ROS) dependent on substrate conditions, O2 concentration, redox state, and activity of the mitochondrial complexes. It is well known that the FADH2-linked substrate succinate induces reverse electron flow to complex I of the electron transport chain and that this process generates superoxide (O2•Ϫ ); these effects are blocked by the complex I blocker rotenone. We demonstrated recently that succinate ϩ rotenone-dependent H 2O2 production in isolated mitochondria increased mildly on activation of the putative big mitochondrial Ca 2ϩ -sensitive K ϩ channel (mtBKCa) by low concentrations of 1,3-dihydro-1-[2-hydroxy-5-(trifluoromethyl)phenyl]-5-(trifluoromethyl)-2H-benzimidazol-2-one (NS-1619). In the present study we examined effects of NS-1619 on mitochondrial O 2 consumption, membrane potential (⌬⌿ m), H2O2 release rates, and redox state in isolated guinea pig heart mitochondria respiring on succinate but without rotenone. NS-1619 (30 M) increased state 2 and state 4 respiration by 26 Ϯ 4% and 14 Ϯ 4%, respectively; this increase was abolished by the BK Ca channel blocker paxilline (5 M). Paxilline alone had no effect on respiration. NS-1619 did not alter ⌬⌿ m or redox state but decreased H 2O2 production by 73% vs. control; this effect was incompletely inhibited by paxilline. We conclude that under substrate conditions that allow reverse electron flow, matrix K ϩ influx through mtBKCa channels reduces mitochondrial H2O2 production by accelerating forward electron flow. Our prior study showed that NS-1619 induced an increase in H 2O2 production with blocked reverse electron flow. The present results suggest that NS-1619-induced matrix K ϩ influx increases forward electron flow despite the high reverse electron flow, and emphasize the importance of substrate conditions on interpretation of effects on mitochondrial bioenergetics.

Mitochondrial depolarization underlies delay in permeability transition by preconditioning with isoflurane: roles of ROS and Ca2+

“…It is suggested that mitochondrial K ϩ channels, often identified as effectors of protection by preconditioning, could be these mediators (10,27,32,34). However, their role in this mechanism is questionable since their activation has been correlated with both increase and decrease in ROS production (20,49). These controversial findings may result from differential behavior of these channels depending on the metabolic state of the cell (44).…”

Ca²⁺-Activated K⁺ Channels as Therapeutic Targets for Myocardial and Vascular Protection

“…Following IR injury Heinen et al demonstrated more efficient forward electron flow with NS1619 treatment. 44 In addition, multiple groups have documented increased respiration following BKCa activation using NS1619 and NS11021. 41, 46 However, in endothelial cells, iberiotoxinsensitive NS1619-induced respiration was associated with increased non-phosphorylating respiration or proton leak.…”

“…40 NS1619 was also shown to attenuate ROS production in isolated mitochondria under conditions to stimulate reverse electron flow. 44 The overall consensus is that activation of the BKCa channel limits ROS following IR, and that this may be one of the major cardioprotective benefits of channel activation.…”

Redox Control of Cardiac Excitability

“…The figure was created by adapting information from (176,195,255), respectively. generation from the cardiac mitochondria, the synthesis of H 2 O 2 from the isolated mitochondria from hearts, or mitochondria in intact cardiac cells, can be determined in a fluorescence-based assay (106). Within these fluorescence-based assays, inhibition of the mitochondrial respiratory chain at any stage causes generation of large amounts of ROS (Fig.…”